A chemical genomics approach toward understanding the global functions of the target of rapamycin protein (TOR)

Abstract

The target of rapamycin protein (TOR) is a highly conserved ataxia telangiectasia-related protein kinase essential for cell growth. Emerging evidence indicates that TOR signaling is highly complex and is involved in a variety of cellular processes. To understand its general functions, we took a chemical genomics approach to explore the genetic interaction between TOR and other yeast genes on a genomic scale. In this study, the rapamycin sensitivity of individual deletion mutants generated by the Saccharomyces Genome Deletion Project was systematically measured. Our results provide a global view of the rapamycin-sensitive functions of TOR. In contrast to conventional genetic analysis, this approach offers a simple and thorough analysis of genetic interaction on a genomic scale and measures genetic interaction at different possible levels. It can be used to study the functions of other drug targets and to identify novel protein components of a conserved core biological process such as DNA damage checkpoint/repair that is interfered with by a cell-permeable chemical compound.

Figure 1

Rapamycin-sensitivity assay for yeast deletion strains. (a) A simplified model for genetic interaction between TOR and other components in the rapamycin-sensitive pathways. (b) The rapamycin-sensitivity assay for yeast deletion mutants of genes known to play a role in rapamycin-sensitive signaling. Wild type yeast (WT), RR strain (TOR1^RR), expressing Tor1p(S1972I) or strains with deletion of GLN3, RRD2, BMH2, URE2, or TOR1 were streaked onto YPD, YPD+25 nM rapamycin, or YPD+25 nM FK506 plates and incubated at 30°C. (c) Scores for rapamycin-sensitivity for strains in b.

Figure 2

Rapamycin hypersensitivity of RH mutants is because of genetic interaction of RH genes with TOR. Selective RH mutants from each functional group carrying pYDF80 expressing RR TOR1(S1972I) or a control plasmid were replica plated onto YPD or YPD+25 nM rapamycin and incubated at 30°C.